1. Field of the Invention
This invention relates to the field of biological cell culture equipment and more particularly to devices for culturing and transport of cells. The present invention involves devices that can be used to support a film, membrane or sponge for the purpose of isolation, expansion, transportation and/or transplantation of cells.
2. Description of Related Art
Cell culture is a process of growing and maintaining cells under laboratory conditions. This process involves isolating the cells from animals or plants, and growing and maintaining the living cells in laboratory devices using artificial culture media to support their basic biological functions. Cell cultures are derived from either primary tissue explants or from cell suspensions. Depending on the nature of the isolated cells, the cells can be either of the adherent or the suspension type. Various methods are known in the art for growing cells in both large scale and small scale cultures.
Various cell culture devices are commercially available in several shapes and designs, the most widely used conventional forms being culture flasks and dishes. The flasks are provided with caps, while the dishes are available as individual or multi well dishes, and are provided with lids.
Typically available individual culture dishes include a base for containing the cultures and a lid to cover the container. The primary purpose of the culture dish is to support cell growth. For practicing cell culture, cells isolated from the tissues are seeded in the culture device along with the nutrient media to nourish the cells, followed by incubation in a controlled. environment. A transparent material like polystyrene is preferred in the manufacture of cell culture dishes to enable visualization of the cultured cells under a microscope.
Conventional cell culture dishes require protection against microbial contamination for successful cell growth, since contamination is one of the principal challenges to successful cell and tissue culturing techniques. Culture dish contamination is typically minimized by using sterilized dishes. The lid serves as a barrier to airborne microbes and prevents evaporation of the media. Conventional cell culture plate lids, however, serve only as a barrier to microbial contamination and do not provide a tight seal that prevents spillage of the contents during transport of the cell cultures.
Specific cells are cultured in dishes under sterile conditions to a desired state of growth for specialized purposes as in the case of medical, scientific or industrial requirements. Following growth, however, these cells have to be removed from their growth environment and shipped to the required end users. For transport of the cultured cells, the cells have to be separated from the dish using enzymes like trypsin, placed within cryoprotectants and shipped under liquid nitrogen. After the cells arrive, they must be re-cultured to maintain their viability. In addition to the time and resources required in re-culturing the cells, this necessity results in significant delays while waiting for the cells to grow to the desired state. The hostile transport conditions also result in considerable loss in viability of cells as well as high shipping costs.
Currently, cell based therapies are being practiced at least in the areas of skin, cornea, cartilage, etc., and the use of cultured cells for medical applications is growing rapidly. There is a growing need, therefore, for an efficient method of delivering cells to the site of transplantation. Cells for use in transplantation therapies can be grown either as monolayers on suitable membranes as described in U.S. Pat. No. 5,693,332 and/or as differentiated into multilayers as published in U.S. Publ. No. 2003/0208266A1 (each incorporated herein by reference in their entireties) before transplantation, or the cells can be cultured on foams or sponges, which can then be transplanted. The important factors involved in membrane-based cell delivery systems include, attachment, viability, cell density, and control over differentiation. The delicate relationship between the cell layers and the membrane can be disturbed during shipment due to movement shear, tear and/or other mechanical damage to the individual cells and their contacts with each other, thus destroying confluence of the monolayer and rendering the cells useless. Although the cells can be cultured on membranes in culture dishes, their transport in the same dishes is not feasible.
Culture flasks were developed to address the problem of contamination in cell culture dishes. Conventional flasks typically have a culture chamber, a small tubular opening located at one end of the flask, and a corresponding closure. The principal designs for various culture flasks are described in U.S. Pat. Nos. 4,334,028, 4,851,351 and 5,398,837 (each incorporated herein by reference in their entirety). As these culture flasks require a relatively large amount of expensive media in order to be filled completely, they are not cost effective. Conventional flasks are also not suitable for the growth of cells for transplantation, as a film or membrane cannot be placed inside the flask to grow cells for transplantation.
Further to dishes and flasks, roller bottles have also been developed for cell culture. Roller bottles offer a larger surface area for cell attachment and growth. Due to the presence of high shear forces associated with these roller bottles, however, this technology is suitable only for those cells that are capable of remaining adhered to the wall of the roller bottle. Also the maintenance of cell cultures in roller bottles for longer periods poses a challenge due to a constant high shear force environment and possible contamination. U.S. Pat. No. 3,450,598 (incorporated herein by reference in its entirety) involves attaching and growing cells on the interior surface of plastic or glass roller tubes and bottles. Other aspects of roller bottle technology are described in U.S. Pat. Nos. 5,527,705 and 4,962,033 (each incorporated herein by reference in their entirety). Further improvements to roller bottles with increased surface areas for cell attachment are described in U.S. Pat. No. 5,010,013 (incorporated herein by reference in its entirety), wherein corrugated channels are added to the interior surface area of the roller bottle. Roller bottles still suffer various inefficiencies, however, including not providing sufficient aeration to the cells, thus diminishing their viability.
More recently, the use of hollow fibers or capillaries has been disclosed as a support matrix for the propagation of cells. This technology is described in U.S. Pat. Nos. 3,821,087, 3,883,393, 3,997,396, 4,184,822, 4,200,689, 4,206,015, 4,220,725, and 4,391,912 (each incorporated herein by reference in their entirety), wherein various apparatus and methods for the in vitro growth of cells on semi-permeable tubular membranes or capillaries are disclosed. The cells are initially allowed to settle onto the surfaces of the capillary walls in a nutrient medium. The nutrients diffuse from the perfusing medium through the capillary walls and are utilized by the cells. Cell products diffuse from the cells through the capillary walls and into the perfusate, from which cell products may be recovered. Unfortunately, nutrient media flow through the hollow capillaries prevents complete penetration of the capillary bundle by the cells and the resultant undesirable gradient of the medium flow leads to incomplete utilization of the available capillary surface for cell attachment. This results in the cells becoming unevenly distributed along the surface. Furthermore, these devices require high media circulation rates to supply adequate oxygen to the cells, and these devices have the additional disadvantages of being mechanically complex, difficult to assemble, and unduly large.
Considering the state of the art, there is a continuing need for an efficient and economical device for culturing, expansion and transport of cell cultures, and it would be beneficial to provide a device suitable for culturing and transporting cell monolayers attached to permeable membranes that are ready to use upon delivery.